Volume 19 Supplement 1

Proceedings of the 1st International Conference of Collaborative Research Center 974: Liver Damage and Regeneration

Open Access

Control and consequences of IL-6 receptor ectodomain shedding

European Journal of Medical Research201419(Suppl 1):S17

https://doi.org/10.1186/2047-783X-19-S1-S17

Published: 19 June 2014

Background

Interleukin-6 type cytokines are mainly involved in inflammation by controlling differentiation, proliferation, migration, and apoptosis of target cells. A dysfunction of the complex regulatory cytokine network might lead to acute and chronic inflammation, autoimmune diseases or neoplastic disorders. IL-6 deficient mice were found to be resistant to collagen- and antigen-induced arthritis, highlighting the role of IL-6 in chronic inflammation and autoimmune diseases. The IL-6 receptor complex consists of the signal-transducing gp130 receptor and the non-signaling IL-6 receptor (IL-6R) which exists in membrane bound and soluble forms. A soluble form of the human IL-6R (sIL-6R) is mainly generated by limited proteolysis (ectodomain shedding) but also by alternative splicing. IL-6 signaling via the membrane-bound IL-6R and gp130 is called classic signaling, whereas IL-6 signaling via the soluble IL-6R and gp130 is referred to as IL-6 trans-signaling. Gp130 is ubiquitously expressed, whereas the membrane-bound IL-6R is mainly expressed on lymphocytes and hepatocytes. Therefore, IL-6 trans-signaling virtually expands IL-6 signaling to all cells of the body. Using the specific trans-signaling inhibitor soluble gp130 (sgp130), we showed that IL-6R ectodomain shedding which facilitates IL-6 trans-signaling is the crucial step in the development and the progression of chronic inflammatory disorders and inflammation-induced cancer.

Structural requirements of IL-6R ectodomain shedding

Most post-transcriptional modifications such as phosphorylation, which control the fate and activity of proteins, are completely reversible. Proteolysis is exceptional because of its irreversible nature. The proteolytic release of membrane-anchored proteins was also called (ectodomain) shedding, and the responsible proteases were termed “sheddases” [1].

Ectodomain shedding of the IL-6R has two functional consequences. First, loss of membrane-bound IL-6R renders the cell non-responsive towards IL-6 classic signaling, thereby abrogating IL-6 signaling. Paradoxically, the generated soluble IL-6R (sIL-6R) itself is biologically active and can mediate IL-6 trans-signaling on cells expressing gp130. A high concentration of 25–50 ng/ml sIL-6R is found in the serum of healthy humans. Under pathophysiological conditions, sIL-6R levels rise up to three fold. The cellular source of the sIL-6R and the in vivo mechanism of its generation are still unclear. The vast majority (90–99%) of the sIL-6R should originate from ectodomain shedding of the membrane-bound precursor, whereas alternative splicing of the IL-6R mRNA is accounts only for a minor proportion (1–10%) [1].

The sIL-6R is generated by constitutive and induced shedding by the “A disintegrin and metalloproteinases” (ADAM)10 and ADAM17 [1]. Recently, we characterized the structural requirements of IL-6R shedding on the site of the substrate [2]. The IL-6R consists of three extracellular domains, important for efficient exocytosis and IL-6 binding. The extracellular domains are followed by a flexible, 52 amino acids long stalk region, a trans-membrane domain and an intracellular domain which regulate basolateral sorting of the IL-6R [3]. The ADAM17 cleavage site was identified within the stalk region between the amino acids 357Q and 358D [1]. Deletion of 10 amino acids surrounding the ADAM17 cleavage site abrogated ADAM17 shedding of the IL-6R but leaves ADAM10 shedding intact. This suggested that the ADAM10 cleavage site is located at a different site. Interestingly, ADAM proteases do not have defined cleavage consensus sequences, which hinder the identification of novel cleavage sites by substrate sequence analysis. Deletion of 5 additional juxtamembrane located amino acids in the aforementioned delta10 IL-6R variant abolishes also ADAM10-mediated ectodomain shedding of the IL-6R [2], suggesting that the ADAM10 cleavage site is in close proximity to the ADAM17 cleavage site. Importantly, both IL-6R variants (delta10 and delta15) are biologically active. Moreover, we showed that only about 20 of the 52 amino acids of the IL-6R stalk region are needed for biological activity of the IL-6R, which supported a model of the IL-6/IL-6R/gp130 signal transducing complex, in which the gp130 receptor chain has a C-shaped structure after IL-6/IL-6R/gp130 complex formation [2]. In this model, the stalk region is necessary for the correct positioning of the IL-6-binding domains of the IL-6R in the signal transducing receptor complex.

Constitutive shedding is dependent on the level of cellular IL-6R expression

Apart from induced IL-6R shedding, which is activated by various substances and conditions, including phorbol esters such as phorbol-12-myristat-13-acetate (PMA), the Ca2+ ionophor ionomycin, extracellular ATP, low membrane-cholesterol levels and apoptosis [1], the IL-6R is also constitutively released from the cell surface, without obvious cellular stimulation. Here, we demonstrated that cellular senescence and EGF-R stimulation lead to increased IL-6R expression, which was regulated via the mTOR pathway [4]. The simple increase in cell surface expressed IL-6R also led to increased generation of sIL-6R, suggesting that the expression level of the cellular substrate indirectly determines the amount of constitutive shedding. sIL-6R serum levels are increased under various inflammatory conditions and up to now, it is not clear if this is dependent on increased IL-6R expression, induced or constitutive shedding. Or data, however, open up the possibility that both mechanisms contribute to inflammation-induced increased sIL-6R levels.

Conclusions

In recent years, we characterized the biological functions of IL-6 trans-signaling and sgp130, which specifically suppresses overshooting IL-6 trans-signaling activities but to leaves “beneficial” IL-6 classic signaling intact. The biological sgp130Fc is currently tested in phase I clinical trials [5]. In contrast to treatment with neutralizing TNFalpha antibodies, administration of the trans-signaling inhibitor sgp130 did not interfere with protective immune responses after infection with Mycobacterium tuberculosis in mice [3]. Moreover, classical signaling was sufficient for early control of Listeria monocytogenes infection in mice [6]. Blockade of IL-6 trans-signaling might therefore exhibit advantages as compared to the global blockade of IL-6 or TNFalpha by monoclonal antibodies for the treatment of chronic inflammatory diseases.

Declarations

Acknowledgements

The work of JS was funded by a grant from the Deutsche Forschungsgemeinschaft, Bonn, Germany (DFG SCHE 907/2-1).

Authors’ Affiliations

(1)
Institute of Biochemistry and Molecular Biology II, Heinrich Heine University

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Copyright

© Scheller; licensee BioMed Central Ltd. 2014

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.